SURFACE MODIFICATION OF ePTFE AND IMPLANTS USING THE SAME

ABSTRACT

A method for modifying an ePTFE surface by plasma immersion ion implantation includes the steps of providing an ePTFE material in a chamber suitable for plasma treatment; providing a continuous low energy plasma discharge onto the sample; and applying negative high voltage pulses of short duration to form a high energy ion flux from the plasma discharge to generate ions which form free radials on the surface of the ePTFE material without changing the molecular and/or physical structure below the surface to define a modified ePTFE surface. The step of applying the high voltage pulses modifies the surface of the ePTFE without destroying the node and fibril structure of the ePTFE, even when the step of applying the high voltage pulses etches and/or carburizes the surface of the ePTFE. The modified surface may have a depth of about 30 nm to about 500 nm. The ions are dosed onto the ePTFE sample at concentrations or doses from about 10 13  ions/cm 2  to about 10 16  ions/cm 2 .

CROSS-REFERENCE TO RELATED APPLICATIONS

This application is a divisional application of U.S. patent applicationSer. No. 11/227,378, filed Sep. 15, 2005, issued as U.S. Pat. No.7,597,924 on Oct. 6, 2009, which claims the benefit of U.S. ProvisionalApplication No. 60/709,257, filed Aug. 18, 2005, the contents of both ofwhich are incorporated herein by reference.

FIELD OF THE INVENTION

The present invention relates to surface modification of ePTFE andimplants using the same. More particularly, the present inventionrelates to modification of ePTFE by plasma immersion ion implantationand fuctionalization of the modified ePTFE.

BACKGROUND OF THE INVENTION

An intraluminal prosthesis is a medical device used in the treatment ofdiseased bodily lumens. One type of intraluminal prosthesis used in therepair and/or treatment of diseases in various body vessels is a stent.A stent is a generally longitudinal tubular device formed ofbiocompatible material which is useful to open and support variouslumens in the body. For example, stents may be used in the vascularsystem, urogenital tract, esophageal tract, tracheal/bronchial tubes andbile duct, as well as in a variety of other applications in the body.These devices are implanted within the vessel to open and/or reinforcecollapsing or partially occluded sections of the lumen.

Stents generally include an open flexible configuration. Thisconfiguration allows the stent to be inserted through curved vessels.Furthermore, this configuration allows the stent to be configured in aradially compressed state for intraluminal catheter implantation. Onceproperly positioned adjacent the damaged vessel, the stent is radiallyexpanded so as to support and reinforce the vessel. Radial expansion ofthe stent may be accomplished by inflation of a balloon attached to thecatheter or the stent may be of the self-expanding variety which willradially expand once deployed. Tubular shaped structures, which havebeen used as intraluminal vascular stents, have included helically woundcoils which may have undulations or zig-zags therein, slotted stents,ring stents, braided stents and open mesh wire stents, to name a few.Super-elastic materials and shape memory materials have been used toform stents.

A graft, including a shunt, for example, a dialysis shunt, is anothercommonly known type of intraluminal prosthesis which is used to repair,replace or bridge various body vessels. A graft provides a lumen throughwhich fluids, such as blood, may flow. Moreover, a graft is oftenconfigured as being generally impermeable to blood to inhibitsubstantial leakage of blood therethrough. Grafts are typically hollowtubular devices that may be formed of a variety of materials, includingtextile and non-textile materials.

A stent and a graft may be combined into a stent-graft endoprosthesis tocombine the features and advantages of each. For example, tubularcoverings have been provided on the inner and/or outer surfaces ofstents to form stent-grafts. It is often desirable to use a thin-walledgraft or covering in the stent-graft endoprosthesis to minimize theprofile of the endoprosthesis and to maximize the flow of blood throughthe endoprosthesis. In such cases non-textile materials, such aspolymeric tubes or sheets formed into tubes, are often used. Expandedpolytetrafluoroethylene or e-PTFE is one common polymeric material usedas the graft portion or covering of a stent-graft endoprosthesis.

Polytetrafluoroethylene (PTFE) is commonly used for implantable medicaldevices due to its chemical stability, bio-stability and bio-inertness.It is also highly hydrophobic. The mechanically stretched, expanded form(ePTFE) is microscopically porous and possesses the stability andinertness properties of PTFE. The hydrophobicity of PTFE and ePTFE andthe low adsorption of proteins by PTFE and ePTFE were regarded asfavourable characteristics for good performance in vascular vessels.Certain considerations, however, are present with implanted PTFE orePTFE materials, including thrombosis and anastomosis stenosis by intimahyperplasia.

A thrombus is the formation of a solid body composed of elements of theblood, e.g., platelets, fibrin, red blood cells, and leukocytes.Thrombus formation is caused by blood coagulation and platelet adhesionto, and platelet activation on, foreign substances. When this occurs, agraft is occluded by such thrombotic material, which in turn, results indecreased patency for the graft. Accordingly, more stringent selectioncriteria are necessary for small caliber vascular graft materialsbecause the small diameters of these grafts magnify the problem ofdeposition of such thrombotic material on the luminal surfaces of thegraft.

Biologically designed PTFE or ePTFE surfaces, e.g. by heparin coating ofthe graft, can reduce platelet adherence and the intima proliferation.Heparin, however, has the problem of a physiological decay in biologicalactivity and in some cases iatrogenic inactivation with protaminsulfate.

Endothelial cells as the inner lining of blood vessels are known toprovide a hemocompatible surface. ePTFE as such, however, does notsupport endothelialization in vivo. Various attempts have been made toachieve an adherent layer of endothelial cells on the surface. Seedingthe cells in a system with dynamic pressure and flow in vitro has beendescribed as one way to obtain an endothelial lining, which resists theshear stress of physiological blood flow.

Some ways of surface modification of ePTFE, which influence polarity andsurface energy, have been successfully attempted, such as surfacetreatment of ePTFE with energetic ions, either as plasma treatment or asion beam irradiation of ePTFE. For example, amide and amine groups weredeposited onto PTFE and ePTFE materials by radio frequency (RF) glowdischarge plasma treatment of butylamine, and bovine aortic endothelialcells were then seeded on the amide/amine coated materials. See, Tsenget al., “Effects Of Amide And Amine Plasma-Treated ePTFE Vascular GraftsOn Endothelial Cell Lining In An Artificial Circulatory System”, JournalOf Biomedical Materials Research, 1998, Volume 42, pages 188-189. SuchRF plasma treatment is typically done at low energy levels to deposit athin coating, such as 30 to 100 Angstrom thick coating as reported inTseng. While, improved cell adherence and proliferation for RF plasmatreated ePTFE was reported as compared to the non-treated ePTFE, longterm stability of the cell layer and the long term stability of theplasma treatment were not adequate because plasma treatment usually onlyhas a relatively short term effect as the modified molecules tend tomigrate into the bulk of the polymer. Ion beam irradiation has been usedto modify the surface of ePTFE. See, Yotoriyama et al., Ion-BeamIrradiated ePTFE For The Therapy Of Intracranial Aneurysms”, No ShinkeiGeka 2004, 32, pages 471-478; Takahashi et al, “Biocompatibility OfePTFE Modified By Ion Beam Irradiation”, No Shinkei Geka 2004, 32, pages339-344. Yotoriyama and Takahashi report improved cell adhesion to theirradiated ePTFE surfaces. The surfaces were irradiated with argon,helium, krypton and neon ions by ion beam techniques with high ionenergy levels of 150 keV. Such high energy levels, however, werereported in Yotoriyama to destroy the fibrils of the ePTFE.

Thus, there is a need in the art for modifying the surface of ePTFE topromote cell adhesion without the disadvantages of the prior art. Inparticular, there is the need for modifying the surface of ePTFE topromote cell adhesion without destroying the node and fibril structureof the ePTFE.

SUMMARY OF THE INVENTION

In one aspect of the present invention, a method for modifying an ePTFEsurface by plasma immersion ion implantation includes the steps ofproviding an ePTFE material on a sample holder in a chamber suitable forplasma treatment; providing a continuous low energy plasma dischargeonto the sample; and applying short negative high voltage pulses ofshort duration to the sample holder to form a high energy ion flux fromthe plasma discharge and directed to the ePTFE material. The freeradials generated on the surface of the ePTFE material without changingthe molecular and/or physical structure below the surface to define amodified ePTFE surface. The ePTFE material has a node and fibrilstructure, and the step of applying the high voltage pulses modifies thesurface of the ePTFE without destroying the node and fibril structure,even when the step of applying the high voltage pulses etches and/orcarburizes the surface of the ePTFE. The modified surface may have adepth of about 30 nm to about 500 nm. Desirably, the ions are dosed ontothe ePTFE sample at concentrations or doses from about 10¹³ ions/cm² toabout 10¹⁶ ions/cm².

The step of providing the continuous low energy plasma discharge ontothe sample may further include the step of generating the plasmadischarge at a radiofrequency of about 13.56 MHz or about 2.45 GHz.These radiofrequencies are non-limiting, and other radiofrequencies maysuitably be used. The step of providing the continuous low energy plasmadischarge onto the sample, may also further include the step ofproviding a source of gas from which the plasma is generated, whereinthe gas is selected from the group consisting of nitrogen, oxygen, argonand combinations thereof.

The step of applying high voltage pulses for a short period of time toform an ion flux from the plasma discharge may further include the stepof applying voltages from about −0.5 kV to about −40 kV. The step ofapplying high voltage pulses for a short period of time to form an ionflux from the plasma discharge may also further include the step ofapplying voltages from about −0.5 kV to about −20 kV. The step ofapplying high voltage pulses for a short period of time to form an ionflux from the plasma discharge may also further include the step ofapplying voltages at a frequency from 0.2 Hz to 200 Hz. The voltages maybe applied for a duration of about 1 to about 10 microseconds, desirablyfor a duration of about 5 microseconds.

The power to generate the plasma discharge according to the presentinvention may vary from 50 watts to 500 watts. The pressure within thechamber may be reduced to a pressure of about 0.1 Pa to about 1.0 Pa.

The method for modifying ePTFE surfaces may further include the step orsteps of oxidizing at least a portion of the free radials;functionalizing the free radical sites with a spacer molecule ormaterial, wherein the spacer molecule or material may be covalentlybonded to the ePTFE; functionalizing the free radical sites withhydrophilic acrylamide groups; covalently bonding the hydrophilicacrylamide groups to the modified ePTFE surface; functionalizing thefree radical sites with polysaccharide hydroxyethyl starch groups;covalently bonding the polysaccharide hydroxyethyl starch groups to themodified ePTFE surface; covalently bonding a bioactive agent bonded tothe modified ePTFE surface; covalently bonding a bioactive agent bondedto the hydrophilic acrylamide groups that are covalently bonded to themodified ePTFE surface; covalently bonding a bioactive agent bonded tothe polysaccharide hydroxyethyl starch groups that are covalently bondedto the modified ePTFE surface; and combinations thereof. Usefulbioactive agents include anti-thrombogenic agents (such as heparin,heparin derivatives, hirudin, acetylsalicylic acid, urokinase, and PPack(dextrophenylalanine proline arginine chloromethylketone);anti-proliferative agents (such as enoxaprin, angiopeptin, or monoclonalantibodies capable of blocking smooth muscle cell proliferation,hirudin, and acetylsalicylic acid); anti-inflammatory agents (such asdexamethasone, prednisolone, corticosterone, budesonide, estrogen,sulfasalazine, and mesalamine);antineoplastic/antiproliferative/anti-mitotic agents (such aspaclitaxel, 5-fluorouracil, cisplatin, vinblastine, vincristine,epothilones, endostatin, angiostatin and thymidine kinase inhibitors);anesthetic agents (such as lidocaine, bupivacaine, and ropivacaine);anti-coagulants (such as D-Phe-Pro-Arg chloromethyl keton, heparin,antithrombin compounds, platelet receptor antagonists, anti-thrombinantibodies, anti-platelet receptor antibodies, aspirin, prostaglandininhibitors, platelet inhibitors and tick antiplatelet peptides);vascular cell growth promotors (such as growth factor inhibitors, growthfactor receptor antagonists, transcriptional activators, andtranslational promotors); vascular cell growth inhibitors (such asgrowth factor inhibitors, growth factor receptor antagonists,transcriptional repressors, translational repressors, replicationinhibitors, inhibitory antibodies, antibodies directed against growthfactors, bifunctional molecules consisting of a growth factor and acytotoxin, bifunctional molecules consisting of an antibody and acytotoxin); cholesterol-lowering agents; vasodilating agents; agentswhich interfere with endogenous vasoactive mechanisms; adhesion factors(such as RGD sequence containing compounds, lysine, poly-L-lysine,antibodies against endothelial cell markers and/or their precursorcells/stem cells, and elastin); and combinations thereof.

The ePTFE sample modified by the methods of the present invention may bean implantable medical device, for example a vascular or non-vasculardevice. Useful devices include, without limitation, a vascular ornon-vascular graft or shunt, a vascular or non-vascular stent, avascular or non-vascular stent-graft, a patch, such as a patch useful inherniorraphy or craniosurgery, a material or sheet for dura replacement,and the like.

In another aspect of the present invention, an implantable medicaldevice is provided. The device includes ePTFE having a surface modifiedby plasma immersion ion implantation; and polysaccharide hydroxyethylstarch groups covalently bonded to the modified surface. The implantablemedical device may further include a bioactive agent bonded to thepolysaccharide hydroxyethyl starch groups. Useful bioactive agentsinclude anti-thrombogenic agents (such as heparin, heparin derivatives,hirudin, acetylsalicylic acid, urokinase, and PPack (dextrophenylalanineproline arginine chloromethylketone); anti-proliferative agents (such asenoxaprin, angiopeptin, or monoclonal antibodies capable of blockingsmooth muscle cell proliferation, hirudin, and acetylsalicylic acid);anti-inflammatory agents (such as dexamethasone, prednisolone,corticosterone, budesonide, estrogen, sulfasalazine, and mesalamine);antineoplastic/antiproliferative/anti-mitotic agents (such aspaclitaxel, 5-fluorouracil, cisplatin, vinblastine, vincristine,epothilones, endostatin, angiostatin and thymidine kinase inhibitors);anesthetic agents (such as lidocaine, bupivacaine, and ropivacaine);anti-coagulants (such as D-Phe-Pro-Arg chloromethyl keton, heparin,antithrombin compounds, platelet receptor antagonists, anti-thrombinantibodies, anti-platelet receptor antibodies, aspirin, prostaglandininhibitors, platelet inhibitors and tick antiplatelet peptides);vascular cell growth promotors (such as growth factor inhibitors, growthfactor receptor antagonists, transcriptional activators, andtranslational promotors); vascular cell growth inhibitors (such asgrowth factor inhibitors, growth factor receptor antagonists,transcriptional repressors, translational repressors, replicationinhibitors, inhibitory antibodies, antibodies directed against growthfactors, bifunctional molecules consisting of a growth factor and acytotoxin, bifunctional molecules consisting of an antibody and acytotoxin); cholesterol-lowering agents; vasodilating agents; agentswhich interfere with endogenous vasoactive mechanisms; adhesion factors(such as RGD sequence containing compounds, lysine, poly-L-lysine,antibodies against endothelial cell markers and/or their precursorcells/stem cells, and elastin); and combinations thereof. Theimplantable medical device may be a graft, including a vascular graft,or a stent-graft, including a vascular stent-graft.

In another aspect of the present invention, an implantable medicaldevice is provided in which the device includes ePTFE having a surfacemodified by plasma immersion ion implantation; and hydrophilicacrylamide groups covalently bonded to the modified surface. Theimplantable medical device may further include a bioactive agent bonded,such as one or more of the above-described agents, to the hydrophilicacrylamide groups. The implantable medical device may be a vascular ornon-vascular graft or shunt, a vascular or non-vascular stent, avascular or non-vascular stent-graft, a patch, such as a patch useful inhemiorraphy or craniosurgery, a material or sheet for dura replacement,and the like.

In another aspect of the present invention, an implanted, surfacemodified ePTFE graft includes ePTFE having a node and fibril structureand having a carburized surface formed by plasma immersion ionimplantation without destroying the node and fibril structure and havingcellular material attached to the fibrils or attached to functionalgroups covalently bonded to the fibrils and substantially coveringand/or filling the nodes. The carburized surface desirably has a depthof about 30 nm to about 500 nm.

In another aspect of the present invention a surface modified ePTFEincludes ePTFE having a node and fibril structure and having acarburized surface formed by plasma immersion ion implantation withoutdestroying the node and fibril structure and having seed cells and/orprotein material attached to the fibrils and/or attached to spacergroups, preferably hydrophilic acrylamide groups and/or polysaccharidehydroxyethyl starch groups, covalently bonded to the fibril. Useful, butnon-limiting, seed cells include epithelial cells (e.g., keratinocytes,hepatocytes), neurons, glial cells, astrocytes, podocytes, mammaryepithelial cells, islet cells, endothelial cells (e.g., aortic,capillary and vein endothelial cells), and mesenchymal cells (e.g.,adipocytes, dermal fibroblasts, mesothelial cells, osteoblasts), smoothmuscle cells, striated muscle cells, ligament fibroblasts, tendonfibroblasts, adult fibroblasts, fibrocytes, chondrocytes, osteocytes,stem cells, genetically modified cells, immunologically masked cells,combinations thereof, and the like. Useful protein material includesfibronectin, laminin, vitronectin, tenascin, entactin, thrombospondin,elastin, gelatin, collagen, fibrillin, merosin, anchorin, chondronectin,link protein, bone sialoprotein, osteocalcin, osteopontin, epinectin,hyaluronectin, undulin, epiligrin, kalinin, combinations thereof and thelike. Such materials may be useful as a scaffold for seeding cells,i.e., in vitro tissue engineering. The carburized surface desirably hasa depth of about 30 nm to about 500 nm.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 is a perspective view of a graft according to the presentinvention.

FIG. 2 is a perspective view of a stent-graft according to the presentinvention.

FIG. 3 is a schematic depiction of a system for plasma immersion ionimplantation according to the present invention.

FIGS. 4A and 4B graphically depict plasma densities for nitrogen plasmadischarge varying with the pressure of the working gas and the plasmapower according to the present invention.

FIG. 5 graphically depicts FTIR spectra of LDPE after plasma immersionion implantation according to the present invention.

FIG. 6 graphically depicts FTIR ATR spectra of PTFE after plasmaimmersion ion implantation energy according to the present invention.

FIG. 7 graphically depicts FTIR ATR spectra of PTFE after plasmaimmersion ion implantation according to the present invention.

FIG. 8 graphically depicts FTIR ATR spectral of ePTFE samples afterplasma immersion ion implantation treatment and post-treatment byacrylamide and polysaccharide hydroxyethyl starch according to thepresent invention.

FIG. 9 graphically depicts FTIR ATR spectra of PTFE after plasmaimmersion ion implantation treatment and post-treatment by acrylamideand by polysaccharide hydroxyethyl starch according to the presentinvention.

FIG. 10 graphically depicts element concentration in PTFE surface independence on plasma immersion ion implantation dose treatment accordingto the present invention.

FIG. 11 depicts increased roughness of the PTFE surface modified byplasma immersion ion implantation according to the present invention.

FIG. 12 graphically depicts contact angles of water on ePTFE, PTFE, andLDPE after plasma immersion ion implantation treatment post-treatmentaccording to the present invention.

FIGS. 13A and 13B graphically depict ageing kinetics of ePTFE and PTFEaccording to the present invention.

FIGS. 14A and 14B graphically depict accelerated ageing kinetics ofePTFE and PTFE according to the present invention according to thepresent invention.

FIGS. 15A through 15D depict morphology of GM7373 cells after exposureto extracts of ePTFE according to the present invention.

FIG. 16 graphically depicts cell count or activity of GM7373 cells afterexposure to with extracts of untreated and treated ePTFE according tothe present invention.

FIG. 17 graphically depicts cell count or activity of GM7373 cells afterexposure to with extracts of untreated and treated ePTFE and controlmaterial according to the present invention.

FIG. 18 graphically depicts cell count or activity of repeatedexperiments on a subset of samples from FIG. 17, normalized to thecontrol material according to the present invention.

DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENT

The present invention is directed to ion implantation into ePTFE byplasma immersion ion implantation (Pill) of nitrogen, oxygen and/orargon ions and a subsequent chemical functionalization of the surface.

Plasma immersion ion implantation modifies the surface modification of apolymer, such as ePTFE, by the penetration of high energy ions into thepolymer, cascades of collisions with atoms of macromolecules and thetransfer of the kinetic energy of the penetrating ion to atoms andelectrons of the polymer macromolecules. The transferred energy is highenough to break chemical bondings in the macromolecule and releasedatoms and electrons fly with high kinetic energy causing new collisionswith nearest macromolecules. As result, chemical breaking of chemicalbondings, ionization, formation of free radicals, electron and phononexcitation of macromolecules occur. The area of such strong structuralchanges of polymer is named an ion track and the size of an ion trackdepends on ion energy, kind of ion and polymer. These reactions occur inthe first stage of the ion beam implantation into polymers during 10⁻⁹to 10⁻⁶ seconds.

After ion penetration, the track field of polymer has a very highconcentration of free radicals, ionized and highly excited parts ofmacromolecules, which induce a number of chemical reactions in thisdestructed field of polymer. The products are amorphous carbon, aromaticcondensed structures, stable and semi-stable free radical structures.The free radicals cause chain reactions of hydrogen breaking frominitial macromolecules and the modifications range significantly deeperthan the track of ions. As used herein the term “carburizing” andvariants thereof refers to the formation of amorphous carbon, aromaticcondensed structures, stable and semi-stable free radical structuresinto a polymer surface, such as ePTFE. The duration of the second stageis much longer than the first stage of structure transformations. Theproperties of polymer surface after ion beam implantation are mostlyrelated to this second stage. During the second structure transformationstage in presence of air the free radical reactions of the modifiedpolymer layer are involve atmospheric oxygen and stableoxygen-containing groups appear in the polymer. These structuretransformations in polymer surface layer can be used for differentapplications including medical devices. Desirably, the carburizedsurface has a depth of about 30 nm to about 500 nm, more desirably fromabout 50 nm to 150 nm, preferably about 100 nm.

In plasma immersion ion implantation, a sample, such as a polymericsample, for example ePTFE, is placed on a holder in continuous lowenergy plasma discharge and high voltage pulses are applied for a shorttime for the formation of an ion flux from the plasma cloud.

While polymer surfaces have been functionalized by plasma treatment. Incontrast to plasma immersion ion implantation, plasma treatment usuallyonly has a relatively short term effect, because the modified moleculestend to migrate into the bulk of the polymer. With plasma immersion ionimplantation, however, the polymer, for example ePTFE is treated withhigher energetic ions, which induce carburization of the surface.Carburization of the polymeric surface prevents this surface fromremodelling, i.e., migration into the polymer bulk.

Ion implantation by plasma immersion ion implantation into polymers maylead to the formation of dangling bondings and free radicals in thepolymer surface. Such free radicals are potentially toxic for adherentcells, even though no such effect was with the methods of the presentinvention as described in the examples below. To reduce the potentialtoxic effects, these bondings may be saturated prior to implantation ofthe ePTFE into a bodily lumen of a patient. In one aspect of the presentinvention, these bondings may be saturated with the highly hydrophilicacrylamide. Useful acrylamides have the structure of

The highly hydrophilic acrylamide covalently bonds to the polymersurface and, due to chemical reaction with active groups on the polymersurface, polyacrylamide forms, as follows:

Free monomers of toxic substances, if any, may then be washed out.

Alternatively to acrylamide, modified polysaccharide hydroxyethyl starch(HAES) may be used to saturate the free radicals on the polymer surface.A useful modified polysaccharide hydroxyethyl starch includes thosematerials having the structural formula of

Useful modified polysaccharide hydroxyethyl starches include thosehaving an average non-limiting molecular weight of about 200 andsubstitution rate from about 0.40 to about 0.55, desirably about 0.5.Desirably, the HAES may have a molecular weight from about 150 to about300, preferably about 200. HAES has been used in clinical application asplasma expanders for the treatment of hypovolaemia and shock. Due to thesubstitution, the HAES is more soluble in water than ordinary starchesand more resistant against degradation. In solution it tends to inhibithaemostatic processes by interaction with clotting factors and bloodplatelets. Endothelial cells also internalize the free form bypinocytosis and eliminate only about 50% of it. The expression ofactivation markers is not modulated by this, but HAES directly inhibitsthe interaction with polymorphonuclear neutrophils.

A method for modifying an ePTFE surface by plasma immersion ionimplantation according to the present invention includes the steps ofproviding an ePTFE material in a chamber suitable for plasma treatment;providing a continuous low energy plasma discharge onto the sample; andapplying high voltage pulses for a short period of time to form a highenergy ion flux from the plasma discharge to generate ions which formfree radials on the surface of the ePTFE material without changing themolecular and/or physical structure below the surface to define amodified ePTFE surface. The step of applying the high voltage pulses iscontrolled to modify the surface of the ePTFE without destroying thenode and fibril structure of the ePTFE. The energy and frequency of thevoltage pulses are also controlled to etch and/or carburize the surfaceof the ePTFE without destroying the node and fibril structure. Thecontinuous low energy plasma discharge is provided by generating theplasma discharge at a radiofrequency of about 13.56 MHz or about 2.45GHz. These radiofrequencies are non-limiting, and other radiofrequenciesmay suitably be used. The source of gas from which the plasma isgenerated may include nitrogen, oxygen, argon and combinations thereof.Desirably, these gaseous ions are used in the methods of the presentinvention to carburize the surface of the ePTFE as compared to prior artmethods where mere coatings of molecules were deposited onto ePTFEsurfaces.

Short term high voltage pulses from about −0.5 kV to about −40 kV areapplied to the sample or the sample holder to accelerate ions from theplasma toward the sample. Voltages from about −0.5 kV to about −30 kV,from about −0.5 kV to about −20 kV, from about −5 kV to about −40 kV,from about −10 kV to about −30 kV, and from about −20 kV to about −30 kVare also useful. The voltages may be applied at a frequency from about0.2 Hz to about 200 Hz. The voltages may also be applied for a durationof about 1 to about 10 microseconds, desirably about 5 microseconds.

Power to generate to the plasma discharge was from about 50 watts toabout 500 watts, desirably from about 50 to about 400 watts. With plasmaimmersion ion implantation, the chamber operates at a reduced, butnon-limiting, pressure of about 0.1 to about 1.0 Pa. Desirably, ions aredosed onto the ePTFE surface at about 10¹³ to about 10¹⁶ ions/cm².

Such methods carburize the ePTFE surface. After carburization of theePTFE surface, the free radicals on the ePTFE, or a portion of the freeradicals on the ePTFE surface, are desirably oxidized by exposing theePTFE material to an oxidizing environment, for example exposure to air.

The free radial sites or the oxidized free radical sites may befunctionalized with a spacer molecule or material. In contrast to thecoating methods of the prior art, the spacer molecule or material iscovalently bonded to the ePTFE. Useful, but non-limiting, examples ofspacer molecules or materials include the above-described hydrophilicacrylamide groups, polysaccharide hydroxyethyl starch groups andcombinations thereof.

Moreover, a bioactive agent may be covalently bonded to the modifiedePTFE surface or to the spacer molecules, i.e., hydrophilic acrylamidegroups, polysaccharide hydroxyethyl starch groups and combinationsthereof. Useful, but non-limiting, bioactive agents includeanti-thrombogenic agents (such as heparin, heparin derivatives, hirudin,acetylsalicylic acid, urokinase, and PPack (dextrophenylalanine prolinearginine chloromethylketone); anti-proliferative agents (such asenoxaprin, angiopeptin, or monoclonal antibodies capable of blockingsmooth muscle cell proliferation, hirudin, and acetylsalicylic acid);anti-inflammatory agents (such as dexamethasone, prednisolone,corticosterone, budesonide, estrogen, sulfasalazine, and mesalamine);antineoplastic/antiproliferative/anti-mitotic agents (such aspaclitaxel, 5-fluorouracil, cisplatin, vinblastine, vincristine,epothilones, endostatin, angiostatin and thymidine kinase inhibitors);anesthetic agents (such as lidocaine, bupivacaine, and ropivacaine);anti-coagulants (such as D-Phe-Pro-Arg chloromethyl keton, heparin,antithrombin compounds, platelet receptor antagonists, anti-thrombinantibodies, anti-platelet receptor antibodies, aspirin, prostaglandininhibitors, platelet inhibitors and tick antiplatelet peptides);vascular cell growth promotors (such as growth factor inhibitors, growthfactor receptor antagonists, transcriptional activators, andtranslational promotors); vascular cell growth inhibitors (such asgrowth factor inhibitors, growth factor receptor antagonists,transcriptional repressors, translational repressors, replicationinhibitors, inhibitory antibodies, antibodies directed against growthfactors, bifunctional molecules consisting of a growth factor and acytotoxin, bifunctional molecules consisting of an antibody and acytotoxin); cholesterol-lowering agents; vasodilating agents; agentswhich interfere with endogenous vasoactive mechanisms; adhesion factors(such as RGD sequence containing compounds, lysine, poly-L-lysine,antibodies against endothelial cell markers and/or their precursorcells/stem cells, and elastin); and combinations thereof.

The modified and/or functionalized ePTFE materials 12 of the presentinvention are useful as an implantable medical device, for example,vascular graft 10, which is depicted in FIG. 1. Graft 10 is an ePTFE orPTFE containing graft. As depicted in FIG. 1, graft 10 is a hollowtubular structure having opposed open ends 14, 16. The presentinvention, however, is not limited to a single lumen tubular graft. Forexample, grafts of the present invention may have branches, for examplea bifurcated graft, or a varied shaped, for example flared or varyingdiameter, wall portion.

As depicted in FIG. 2, functionalized ePTFE materials 22 of the presentinvention are useful as an implantable medical device, for example,stent-graft 20. Stent-graft 20 includes functionalized ePTFE material22, which may be in the form of a graft covering or otherwise disposedover a stent 28. Typically, the stent-graft 20 is a hollow tubulardevice having opposed open ends 24, 26. The stent 28 includes elongatemembers 30, such as wire strands formed into a hollow tubular structure.

Desirably, the elongate strands or wires 30 are made from nitinol,stainless steel, cobalt-based alloy such as Elgiloy, platinum, gold,titanium, tantalum, niobium, polymeric materials and combinationsthereof. Useful and nonlimiting examples of polymeric stent materialsinclude poly(L-lactide) (PLLA), poly(D,L-lactide) (PLA), poly(glycolide)(PGA), poly(L-lactide-co-D,L-lactide) (PLLA/PLA),poly(L-lactide-co-glycolide) (PLLA/PGA), poly(D,L-lactide-co-glycolide)(PLA/PGA), poly(glycolide-co-trimethylene carbonate) (PGA/PTMC),polydioxanone (PDS), Polycaprolactone (PCL), polyhydroxybutyrate (PHBT),poly(phosphazene) poly(D,L-lactide-co-caprolactone) PLA/PCL),poly(glycolide-co-caprolactone) (PGA/PCL), poly(phosphate ester) and thelike. Wires made from polymeric materials may also include radiopaquematerials, such as metallic-based powders, particulates or pastes whichmay be incorporated into the polymeric material. For example theradiopaque material may be blended with the polymer composition fromwhich the polymeric wire is formed, and subsequently fashioned into thestent as described herein. Alternatively, the radiopaque material may beapplied to the surface of the metal or polymer stent. In eitherembodiment, various radiopaque materials and their salts and derivativesmay be used including, without limitation, bismuth, barium and its saltssuch as barium sulfate, tantalum, tungsten, gold, platinum and titanium,to name a few. Additional useful radiopaque materials may be found inU.S. Pat. No. 6,626,936, which is herein incorporated in its entirely byreference. Metallic complexes useful as radiopaque materials are alsocontemplated. The stent may be selectively made radiopaque at desiredareas along the wire or made be fully radiopaque, depending on thedesired end-product and application. Further, the wires 30 have an innercore of tantalum, gold, platinum, iridium or combination of thereof andan outer member or layer of nitinol to provide a composite wire forimproved radiocapicity or visibility. Desirably, the inner core isplatinum and the outer layer is nitinol. More desirably, the inner coreof platinum represents about at least 10% of the wire based on theoverall cross-sectional percentage. Moreover, nitinol that has not beentreated for shape memory such as by heating, shaping and cooling thenitinol at its martensitic and austenitic phases, is also useful as theouter layer. Further details of such composite wires may be found inU.S. Patent Application Publication 2002/0035396 A1, the contents ofwhich is incorporated herein by reference.

Various stent types and stent constructions may be employed in theinvention as the stent 20. Among the various stents useful include,without limitation, self-expanding stents and balloon expandableextents. The stents may be capable of radially contracting, as well andin this sense can best be described as radially distensible ordeformable. Self-expanding stents include those that have a spring-likeaction which causes the stent to radially expand, or stents which expanddue to the memory properties of the stent material for a particularconfiguration at a certain temperature. Nitinol is one material whichhas the ability to perform well while both in spring-like mode, as wellas in a memory mode based on temperature. Other materials are of coursecontemplated, such as stainless steel, platinum, gold, titanium andother biocompatible metals, as well as polymeric stents. Theconfiguration of the stent may also be chosen from a host of geometries.For example, wire stents can be fastened into a continuous helicalpattern, with or without a wave-like or zig-zag in the wire, to form aradially deformable stent. Individual rings or circular members can belinked together such as by struts, sutures, welding or interlacing orlocking of the rings to form a tubular stent. Tubular stents useful inthe present invention also include those formed by etching or cutting apattern from a tube. Such stents are often referred to as slottedstents. Furthermore, stents may be formed by etching a pattern into amaterial or mold and depositing stent material in the pattern, such asby chemical vapor deposition or the like. Examples of various stentconfigurations are shown in U.S. Pat. No. 4,503,569 to Dotter; U.S. Pat.No. 4,733,665 to Palmaz; U.S. Pat. No. 4,856,561 to Hillstead; U.S. Pat.No. 4,580,568 to Gianturco; U.S. Pat. No. 4,732,152 to Wallsten, U.S.Pat. No. 4,886,062 to Wiktor, and U.S. Pat. No. 5,876,448 to Thompson,all of whose contents are incorporated herein by reference.

A system 40 for immersion ion implantation of samples, such as PTFE orePTFE samples, is depicted in FIG. 3. The system 40 includes a chamber42, a radiofrequency antenna 44, a high voltage electrode 46, a sampleholder 48, a plasma generator 50, a high voltage generator 52, aturbo-molecular pump 54 and a vacuum pump 56, interrelated as shown. Asample (not shown) may be placed on or otherwise secured to the sampleholder 48. The turbo-molecular pump 54 and the vacuum pump 56 are usefulfor controlling the pressure and flow of gas within the chamber 42. Theplasma generator 50 and the radiofrequency antenna 44 are useful forgenerating a continuous low plasma discharge (not shown). The highvoltage electrode 46 and the high voltage generator 52 are useful forgenerating and applying high voltage pulses of short duration to form ahigh energy ion flux from the plasma discharge to generate ion capableof forming free radials on the surface of the sample, i.e., high energyions.

In one aspect of the present invention, a method for modifying an ePTFEsurface by plasma immersion ion implantation includes the steps ofproviding an ePTFE material in a chamber suitable for plasma treatment;providing a continuous low energy plasma discharge onto the sample; andapplying high voltage pulses for a short period of time to form a highenergy ion flux from the plasma discharge to generate ions which formfree radials on the surface of the ePTFE material without changing themolecular and/or physical structure below the surface to define amodified ePTFE surface. The ePTFE material has a node and fibrilstructure, and the step of applying the high voltage pulses modifies thesurface of the ePTFE without destroying the node and fibril structure,even when the step of applying the high voltage pulses etches and/orcarburizes the surface of the ePTFE. The modified surface may have adepth of about 30 nm to about 500 nm. Desirably, the ions are dosed ontothe ePTFE sample at concentrations or doses from about 10¹³ ions/cm² toabout 10¹⁶ ions/cm².

The step of providing the continuous low energy plasma discharge ontothe sample may further include the step of generating the plasmadischarge at a radiofrequency of about 13.56 MHz or about 2.45 GHz. Suchuseful radiofrequencies are non-limiting and other radiofrequencies maysuitably be used. The step of providing the continuous low energy plasmadischarge onto the sample, may also further include the step ofproviding a source of gas from which the plasma is generated, whereinthe gas is selected from the group consisting of nitrogen, oxygen, argonand combinations thereof.

The step of applying high voltage pulses for a short period of time toform an ion flux from the plasma discharge may further include the stepof applying voltages from about −0.5 kV to about −40 kV. The step ofapplying high voltage pulses for a short period of time to form an ionflux from the plasma discharge may also further include the step ofapplying voltages from −0.5 kV to −20 kV. The step of applying highvoltage pulses for a short period of time to form an ion flux from theplasma discharge may also further include the step of applying voltagesat a frequency from 0.2 Hz to 200 Hz. The voltages may be applied for aduration of about 1 to about 10 microseconds, desirably for a durationof about 5 microseconds.

The power to generate the plasma discharge according to the presentinvention may vary from 50 watts to 500 watts. The pressure within thechamber may be reduced to a pressure of about 0.1 Pa to about 1.0 Pa.

The method for modifying ePTFE surfaces may further include the step orsteps of oxidizing at least a portion of the free radials;functionalizing the free radical sites with a spacer molecule ormaterial, wherein the spacer molecule or material is covalently bondedto the ePTFE; functionalizing the free radical sites with hydrophilicacrylamide groups; covalently bonding the hydrophilic acrylamide groupsto the modified ePTFE surface; functionalizing the free radical siteswith polysaccharide hydroxyethyl starch groups; covalently bonding thepolysaccharide hydroxyethyl starch groups to the modified ePTFE surface;covalently bonding a bioactive agent bonded to the modified ePTFEsurface; covalently bonding a bioactive agent bonded to the hydrophilicacrylamide groups that are covalently bonded to the modified ePTFEsurface; covalently bonding a bioactive agent bonded to thepolysaccharide hydroxyethyl starch groups that are covalently bonded tothe modified ePTFE surface; and combinations thereof. Useful bioactiveagents include anti-thrombogenic agents (such as heparin, heparinderivatives, hirudin, acetylsalicylic acid, urokinase, and PPack(dextrophenylalanine proline arginine chloromethylketone);anti-proliferative agents (such as enoxaprin, angiopeptin, or monoclonalantibodies capable of blocking smooth muscle cell proliferation,hirudin, and acetylsalicylic acid); anti-inflammatory agents (such asdexamethasone, prednisolone, corticosterone, budesonide, estrogen,sulfasalazine, and mesalamine);antineoplastic/antiproliferative/anti-mitotic agents (such aspaclitaxel, 5-fluorouracil, cisplatin, vinblastine, vincristine,epothilones, endostatin, angiostatin and thymidine kinase inhibitors);anesthetic agents (such as lidocaine, bupivacaine, and ropivacaine);anti-coagulants (such as D-Phe-Pro-Arg chloromethyl keton, heparin,antithrombin compounds, platelet receptor antagonists, anti-thrombinantibodies, anti-platelet receptor antibodies, aspirin, prostaglandininhibitors, platelet inhibitors and tick antiplatelet peptides);vascular cell growth promotors (such as growth factor inhibitors, growthfactor receptor antagonists, transcriptional activators, andtranslational promotors); vascular cell growth inhibitors (such asgrowth factor inhibitors, growth factor receptor antagonists,transcriptional repressors, translational repressors, replicationinhibitors, inhibitory antibodies, antibodies directed against growthfactors, bifunctional molecules consisting of a growth factor and acytotoxin, bifunctional molecules consisting of an antibody and acytotoxin); cholesterol-lowering agents; vasodilating agents; agentswhich interfere with endogenous vasoactive mechanisms; adhesion factors(such as RGD sequence containing compounds, lysine, poly-L-lysine,antibodies against endothelial cell markers and/or their precursorcells/stem cells, and elastin); and combinations thereof.

The ePTFE sample modified by the methods of the present invention may bean implantable medical device, for example, a vascular or non-vasculargraft or shunt, a vascular or non-vascular stent, a vascular ornon-vascular stent-graft, a patch, such as a patch useful in hemiorraphyor craniosurgery, a material or sheet for dura replacement, and thelike.

In another aspect of the present invention, an implantable medicaldevice is provided. The device includes ePTFE having a surface modifiedby plasma immersion ion implantation; and polysaccharide hydroxyethylstarch groups covalently bonded to the modified surface. The implantablemedical device may further include a bioactive agent bonded to thepolysaccharide hydroxyethyl starch groups. Useful, but non-limiting,bioactive agents include anti-thrombogenic agents (such as heparin,heparin derivatives, hirudin, acetylsalicylic acid, urokinase, and PPack(dextrophenylalanine proline arginine chloromethylketone);anti-proliferative agents (such as enoxaprin, angiopeptin, or monoclonalantibodies capable of blocking smooth muscle cell proliferation,hirudin, and acetylsalicylic acid); anti-inflammatory agents (such asdexamethasone, prednisolone, corticosterone, budesonide, estrogen,sulfasalazine, and mesalamine);antineoplastic/antiproliferative/anti-mitotic agents (such aspaclitaxel, 5-fluorouracil, cisplatin, vinblastine, vincristine,epothilones, endostatin, angiostatin and thymidine kinase inhibitors);anesthetic agents (such as lidocaine, bupivacaine, and ropivacaine);anti-coagulants (such as D-Phe-Pro-Arg chloromethyl keton, heparin,antithrombin compounds, platelet receptor antagonists, anti-thrombinantibodies, anti-platelet receptor antibodies, aspirin, prostaglandininhibitors, platelet inhibitors and tick antiplatelet peptides);vascular cell growth promotors (such as growth factor inhibitors, growthfactor receptor antagonists, transcriptional activators, andtranslational promotors); vascular cell growth inhibitors (such asgrowth factor inhibitors, growth factor receptor antagonists,transcriptional repressors, translational repressors, replicationinhibitors, inhibitory antibodies, antibodies directed against growthfactors, bifunctional molecules consisting of a growth factor and acytotoxin, bifunctional molecules consisting of an antibody and acytotoxin); cholesterol-lowering agents; vasodilating agents; agentswhich interfere with endogenous vasoactive mechanisms; adhesion factors(such as RGD sequence containing compounds, lysine, poly-L-lysine,antibodies against endothelial cell markers and/or their precursorcells/stem cells, and elastin); and combinations thereof. Theimplantable medical device may be a graft, including a vascular graft,or a stent-graft, including a vascular stent-graft.

In another aspect of the present invention, an implantable medicaldevice is provided in which the device includes ePTFE having a surfacemodified by plasma immersion ion implantation; and hydrophilicacrylamide groups covalently bonded to the modified surface. Theimplantable medical device may further include a bioactive agent bonded,such as one or more of the above-described agents, to the hydrophilicacrylamide groups. The implantable medical device may be a vascular ornon-vascular graft or shunt, a vascular or non-vascular stent, avascular or non-vascular stent-graft, a patch, such as a patch useful inherniorraphy or craniosurgery, a material or sheet for dura replacement,and the like.

In another aspect of the present invention, an implanted, surfacemodified ePTFE graft includes ePTFE having a node and fibril structureand having a carburized surface formed by plasma immersion ionimplantation without destroying the node and fibril structure and havingcellular material attached to the fibrils or attached to functionalgroups covalently bonded to the fibrils and substantially coveringand/or filling the nodes. The carburized surface desirably has a depthof about 30 nm to about 500 nm.

In another aspect of the present invention a surface modified ePTFEincludes ePTFE having a node and fibril structure and having acarburized surface formed by plasma immersion ion implantation withoutdestroying the node and fibril structure and having seed cells and/orprotein material attached to the fibrils and/or attached to spacergroups, preferably hydrophilic acrylamide groups and/or polysaccharidehydroxyethyl starch groups, covalently bonded to the fibril. Useful, butnon-limiting, seed cells include epithelial cells (e.g., keratinocytes,hepatocytes), neurons, glial cells, astrocytes, podocytes, mammaryepithelial cells, islet cells, endothelial cells (e.g., aortic,capillary and vein endothelial cells), and mesenchymal cells (e.g.,adipocytes, dermal fibroblasts, mesothelial cells, osteoblasts), smoothmuscle cells, striated muscle cells, ligament fibroblasts, tendonfibroblasts, adult fibroblasts, fibrocytes, chondrocytes, osteocytes,stem cells, genetically modified cells, immunologically masked cells,combinations thereof, and the like. Useful protein material includesfibronectin, laminin, vitronectin, tenascin, entactin, thrombospondin,elastin, gelatin, collagen, fibrillin, merosin, anchorin, chondronectin,link protein, bone sialoprotein, osteocalcin, osteopontin, epinectin,hyaluronectin, undulin, epiligrin, kalinin, combinations thereof and thelike. Such materials may be useful as a scaffold for seeding cells,i.e., in vitro tissue engineering. The carburized surface desirably hasa depth of about 30 nm to about 500 nm.

The following non-limiting examples are intended to further illustratethe present invention.

EXAMPLES

Plasma immersion ion implantation of nitrogen, oxygen and argon ionswith different energies and different regimes of treatment was used forthe modification of ePTFE sheets, PTFE thin films and low-densitypolyethylene (LDPE) films. The PTFE and LDPE samples were used assatellite samples for supporting analysis.

Materials and Methods

Materials

ePTFE provided by Boston Scientific SCIMED was used for plasma immersionion implantation treatment, structure analysis and for cell cultureexperiments. PTFE films of 20 μm and LDPE films of 50 μm were used forplasma immersion ion implantation treatment and structure analysis. ThePTFE and LDPE films were cleaned by alcohol before plasma immersion ionimplantation. The ePTFE samples were not cleaned before plasma immersionion implantation, but the surface of the samples was not touched afterremoving from the packing.

Plasma Immersion Ion Implantation

Most of the modifications were done with the equipment of theForschungszentrum Rossendorf (FZR), Dresden Germany. The pressure ofresidual air was 10⁻³ Pa, working pressure at discharge was 10⁻¹ Pa.Nitrogen, oxygen and argon gases were used for plasma discharge. Plasmawas generated by radiofrequency generator of 13.56 MHz. Plasma power wasregulated in the range of 50-400 W. High voltage pulses were applied tothe sample holder at the stable plasma discharge after 0.5-1 minutesafter plasma start. The high voltage pulses had 5 μs duration; 20 kV, 10kV, 1 kV and 0.5 kV values of peak voltage were used. A pulse repetitionfrequency from 0.2 Hz to 200 Hz was used. The regulation of the pulsefrequency was used to control the temperature during the plasmaimmersion ion implantation treatment. Plasma immersion ion implantationtreatment with doses from 10¹³ to 10¹⁶ ions/cm² was carried out forePTFE, PTFE and LDPE samples.

Additionally, some samples were treated in the Institute of SurfaceModification, Leipzig. In this case only nitrogen ions were used withenergy of 20 keV. The dose of treatment was 10¹³, 10¹⁴, 10¹⁵ and 10¹⁶cm⁻².

The dose of plasma immersion ion implantation treatment was calculatedon direct measurement of plasma density with a Langmuir probe and bycomparison of Fourier-Transform-Infrared (FTIR) spectra of the satellitesamples of LDPE in comparison with well known previous data. By Langmuirprobe the stability of the plasma density was determined and a deviationof dose treatment was estimated as 10% of the average value. Forexample, the behaviour of plasma density for nitrogen plasma dischargein dependence on pressure of working gas and plasma power are presentedin FIGS. 4A and 4B. The plasma density is a function of the chamberpressure and the power used to generate the plasma. The plasma densityincreases with increases of both of these variables.

The list of the plasma immersion ion implantation regimes used forePTFE, PTFE and LDPE treatment is presented in Table 1.

TABLE 1 Chamber Energy Dose Working Used [keV] [ions/cm²] Gas FZR 2010¹³, 10¹⁴, 10¹⁵, 10¹⁶ Nitrogen FZR 10 10¹³, 10¹⁴, 10¹⁵, 10¹⁶ NitrogenFZR 1 10¹³, 10¹⁴, 10¹⁵, 10¹⁶ Nitrogen FZR 0.5 10¹³, 10¹⁴, 10¹⁵, 10¹⁶Nitrogen FZR 20 10¹³, 10¹⁴, 10¹⁵, 10¹⁶ Argon FZR 10 10¹³, 10¹⁴, 10¹⁵,10¹⁶ Argon FZR 1 10¹³, 10¹⁴, 10¹⁵, 10¹⁶ Argon FZR 0.5 10¹³, 10¹⁴, 10¹⁵,10¹⁶ Argon FZR 20 10¹³, 10¹⁴, 10¹⁵, 10¹⁶ Oxygen FZR 10 10¹³, 10¹⁴, 10¹⁵,10¹⁶ Oxygen FZR 1 10¹³, 10¹⁴, 10¹⁵, 10¹⁶ Oxygen FZR 0.5 10¹³, 10¹⁴,10¹⁵, 10¹⁶ Oxygen Leipzig 20 10¹³, 10¹⁴, 10¹⁵, 10¹⁶ Nitrogen

Chemical Post-Treatment

The modified samples were treated by a 10% solution of acrylamide inwater a 10% solution of the modified polysaccharide hydroxyethyl starchin water (HAES sterile 10%, Fresenius AG). The post-treatment was doneimmediately after plasma immersion ion implantation, as well as after 20days and after different time of an accelerating ageing procedure. Thesamples were completely immersed in the solutions for 2 hours at roomtemperature. After the post-treatment the samples were washed withdeionized water and dried on air. The wetting angle measurement of thesesamples was determined at the following day to exclude any influence ofresidual water on surface of the polymers.

FTIR ATR Spectra

FTIR ATR spectra were recorded on Nicolet Magna spectrometer with Ge ATRcrystal and KRS-6 ATR crystal. Number of scans was 100; spectralresolution was 2 cm⁻¹.

Contact Angle Measurement

The measurements of contact angles were done with water and CH₂I₂ dropson a Krüss drop shape device using the sessile drop method. The wettingangle was determined by recalculating the drop geometry of video-imagesand calculating tangents to a baseline.

Accelerating Ageing of Modified ePTFE

Thermal treatment was used for acceleration of ageing processes afterplasma immersion ion implantation in modified ePTFE and PTFE samples.The thermobox provided a temperature of 120° C. in dry air. Thistemperature was selected for ageing test, because of the high stabilityof PTFE up to 350-400° C., the stability of oxygen-containing productsof the surface layer destruction at this temperature but not at highertemperature and a high rate of the free radical quenching at thistemperature. Besides that, clinical sterilization by autoclavation alsois performed at this temperature.

The ageing processes of polymers are very complex and a precisecalculation of the energetic parameters of the ageing processes isimpossible. An estimation of the ageing period was done according to VanHoff's law for temperature dependence of chemical reactions rate. 10° K.increase of temperature represent a duplication of the reaction time.For 100° K. above ambient temperature an accelerated ageing by factor1000 is expected, 45 min represent 1 month.

Cell Culture

A three-step approach was chosen to select an optimal treatment regime.

Short-term cell performance on single regime treated ePTFE versusuntreated ePTFE. This gives some first hints about possibly toxic sideproducts by the ion implantation.

Screening of an array of three ion implantation regimes and two methodsof functionalization

Further studies with samples of few selected parameter sets.

Step (1)

ePTFE discs were mounted in Minusheets (Minucells and Minutissue, BadAbbach) and steam sterilized at 120° C. The bovine aortic endothelialcell line GM7373 (Coriell) was used for the experiments. 4×10⁴ cells in200 μL medium (MEM-Earle supplemented with 10% FBS, 1×MEM vitamins, 2 mMN-Acetyl L-Alanyl L-Glutamine, 1× Amino acids) were seeded directly onthe samples (area 0.58 cm²). They were allowed to adhere for 2 hours atstandard cell culture conditions, then medium was filled up to 1 mL persample.

17 hours after seeding cells of one sample each were fixed in 0.2%paraformaldehyde in PBS and stained with phalloidin-TRITC and DAPI forpolymerized actin of the cytoskeleton (red) and cell nuclei (blue).Images were taken by fluorescent microscopy and digitally overlaid.

The other samples with cell culture 17 hours after seeding were placedin 500 μL MEM Earle w/o phenol red, supplemented with 10% FBS, 10%Alamar Blue™. The extinction of the medium was measured after 4.5 hoursat λ=570 nm and 600 nm against a blank medium without cells. The valueof(E_(blank, 600 nm)−E_(cells, 600 nm))+(E_(cells, 570 nm)−E_(blank, 570 nm))is a measure of the cell activity. It was calibrated with the activityof defined cell numbers. The number of vital cells on the samples wasconcluded from their metabolic equivalent.

Toxicity further was tested in a MEM elution test close to DIN EN ISO10993-5: 1999: An extract of plasma immersion ion implantation treatedePTFE (10 ¹⁶ N_(x) ⁺ cm⁻²), macroscopically 25 cm² treated surface wassteam sterilized at 120° C., 20 min and immersed in 5 mL cell culturemedium (MEM Earle with the supplements) in a vial of polypropylene for46 hours at 37° C. with repeated mixing under sterile conditions. Ascontrols an extract was made of an equivalent amount of untreated ePTFEand medium in an empty vial. Positive controls were medium with CuCl₂ inmedium (1 mM, 250 μM, 62.5 μM, and 15.5 μM).

1 mL of the extracts was added to 2 cm² confluent cell layer of GM7373each. Additionally 1:2, 1:4, and 1:8 dilutions of the extracts withfresh medium were tested. The cells were kept under standard cellculture conditions for three days and inspected for morphologicalchanges daily. At the third day the cell layers were washed once withPBS and the metabolic activity was checked with the chromogenicsubstrate Alamar Blue™ as described above. The test was performed induplicate with the extracts and once with the controls.

Step (2)

Samples as in Table 2 were used. Cells were seeded out as described instep (1). An Alamar Blue™ test was performed at day 1, 3, and 5. Theexperiments were performed in duplicate.

TABLE 2 Energy Dose Label [keV] [N cm⁻²] Functionalization Thermanox ⁽¹⁾— — — untreated — — — 1e14 20 10¹⁴ — 1e15 20 10¹⁵ — 1e16 20 10¹⁶ — 1e14AA 20 10¹⁴ Acrylamide 1e15 AA 20 10¹⁵ Acrylamide 1e16 AA 20 10¹⁶Acrylamide 1e14 HAES 20 10¹⁴ HAES sterile 10% 1e15 HAES 20 10¹⁵ HAESsterile 10% 1e16 HAES 20 10¹⁶ HAES sterile 10% ⁽¹⁾ Thermanox ® (Nunc,Wiesbaden) is a polyethylene terephthalate, treated for cell cultureapplications.

Step (3)

The samples 1e16, 1e16 AA, 1e16 HAES, and 1e14 HAES were selected forfurther investigation besides the cell culture treated Thermanox® anduntreated ePTFE as controls. As used herein, the shorthand of “1e16” andthe like may be used interchangeably with 10¹⁶, or the like.

Cells were seeded out as described above, but at various cell densitiesfor the different repeats. An Alamar Blue™ test was performed on day 1and 3 after seeding. For one set of samples, six hours after seedingcells were fixed with 2% paraformaldehyde and a fluorescent staining wasperformed as described above.

Results

Color Changes of the Samples

The plasma immersion ion implantation treatment induces a change ofcolor of the samples at high doses. LDPE samples get a metallic shade.PTFE and ePTFE samples become gray. A homogeneous distribution of thecolor is an indicator of an inhomogeneous dose distribution on thesample surface. In all cases the dose was homogeneously distributed,excluding some edges where samples are attached by metal screws tosample holder. These parts of the samples are excluded from followinganalysis of the structure and cell experiments.

Molecular Structure of Surface

Plasma immersion ion implantation only modified a thin surface layer ofthe polymers. Therefore no changes of transmission spectra of thetreated samples for all kinds of polymers could be observed.

In FTIR spectra of LDPE after plasma immersion ion implantation thereare some new spectral lines as depicted in FIG. 5. In FIG. 5, the arrayshows the dose increase, i.e., initial sample, 10¹³, 10¹⁴, 5*10¹⁴, 10¹⁵,5*10¹⁵, 10¹⁶ ion/cm², plasma immersion ion implantation of N⁺ ions at 20keV energy. The 1750 cm⁻¹ line corresponds to carbonyl group vibrationsin new oxygen-containing groups, the 1650 cm⁻¹ line corresponds tounsaturated carbon-carbon group vibrations in aromatic and unsaturatedgroups, the field of 1100 cm⁻¹ corresponds to oxygen-containing groupvibrations like ether groups, the lines at 881, 907 and 968 cm⁻¹correspond to vibrations of unsaturated vinyl, vinylidene andcis-vinylene groups. The appearance of such groups is explained byradiation reactions in surface layer under plasma immersion ionimplantation treatment.

FIG. 6 depicts the FTIR ATR spectra of PTFE after plasma immersion ionimplantation with N+ at 20 keV. The arrow indicates the plasma immersionion implantation dose increase: untreated sample, 10¹⁴, 10¹⁵ and 10¹⁶ions/cm². As depicted in FIG. 6, the spectra of PTFE the spectralchanges were less prominent. The lines at 1750 and 1650 cm⁻¹ areobserved in the modified samples with low intensity. These linescorrespond to appearance of oxygen-containing groups and aromaticcondensed structures in PTFE surface layer. The new groups appear due tothe destruction processes of the surface layer and reactions of freeradicals with oxygen of air after plasma immersion ion implantationtreatment. The low intensity of the changes in the FTIR ATR spectra area consequence of significant differences of the depth resolution ofanalysis technique and thickness of the modified layer. The thickness ofthe modified PTFE layer at nitrogen plasma immersion ion implantation at20 keV is about 100 nm, but the depth of ATR layer analysis for GEcrystal equals to 1000-700 nm. Besides that, the sputtering effect ofthe ions for PTFE is much higher than for LDPE. Therefore plasmamodification of PTFE generally does not have a wide application. In thecase of plasma immersion ion implantation a carbonization and oxidationof the surface layer of PTFE were observed.

FIG. 7 depicts FTIR ATR spectra of PTFE after plasma immersion ionimplantation with N+ at 20 keV and at 10¹⁶ ions/cm². The samplesdepicted are as follows: bottom: plasma immersion ion implantationtreated sample; middle: post plasma immersion ion implantation treatmentby acrylamide; top: post plasma immersion ion implantation treatment byHAES. Due to the ion implantation the PTFE surface showed chemicalreactivity. The high activity of free radicals allows a surfacemodification of PTFE and ePTFE with a wide number of substances. Inthese experiments the PTFE surface was treated with an acrylamidesolution and a HAES solution. In FTIR spectra after acrylamidepost-treatment (FIG. 7 middle) there are lines at 2994, 2960, 2924, 2854cm⁻¹ interpreted as stretch vibrations and 1465, 1452 cm⁻¹ interpretedas deformation vibrations of C—H bonds in the acrylamide gel on the PTFEsurface. These lines do not disappear after washing of PTFE in water. Othe untreated reverse side of PTFE these lines could not be observed.After post-treatment of PTFE with HAES (FIG. 7 top) the spectra containthe lines at 2956, 2925, 2855, 1451 cm⁻¹ interpreted as C—H stretch anddeformational vibrations of the crosslinked HAES layer. The line at 1028cm⁻¹ in wind of the strong PTFE line is interpreted as C—O vibrations ofHAES layer. This layer is observed only on the treated side of PTFE andis stable after washing of the ePTFE.

As depicted in FIG. 8, the same spectral changes were observed for ePTFEsamples after plasma immersion ion implantation treatment andpost-treatment by acrylamide and HAES. FIG. 8 depicts the FTIR ATRspectra of ePTFE, 20 keV, N+, 10 ¹⁶ ions/cm²: a —HAES post-treatment,b—acrylamide post-treatment.

FIG. 9 depicts the FTIR ATR spectra of PTFE, plasma immersion ionimplantation treatment, 20 keV, N+, by arrow: 10¹⁴, 10¹⁵, 10¹⁶ ions/cm²and post-treatment by acrylamide, 10¹⁴, 10¹⁵, 10¹⁶ ions/cm² andpost-treatment by HAES. FIG. 9 shows that there was a dose dependence ofthe treatment dose and the amount of crosslinked acrylamide and HAES.

FIG. 10 depicts element concentration in PTFE surface in dependence onplasma immersion ion implantation dose treatment (N⁺, 20 keV). Curveswere generated by XPS data. Strong chemical changes of PTFE could beobserved by XPS, which were used for element contamination analysis ofthe surface layer of PTFE after plasma immersion ion implantation. Inmodified PTFE the contamination of fluorine atoms was significantlylower then in initial. The relative concentration of carbon atomsincreased. The appearance of nitrogen and oxygen atoms was observed.However, there was no strong dependence of the atomic concentrations ofelements on dose of plasma immersion ion implantation treatment. Even atlow dose, the surface layer of PTFE contained a significant amount ofoxygen and nitrogen atoms. At high dose, the effect of etching becamesignificant and the concentration of fluorine atoms started to increaseagain, parallel with a decrease of oxygen and nitrogen. This indicatesthat the etching effect becomes significant at high dose treatment.

The etching process also could be demonstrated morphologically on thePTFE surface. As depicted in FIG. 11, atomic force microscopy (AFM)shows an increased roughness of the PTFE surface modified by plasmaimmersion ion implantation with N⁺ at 20 keV. Although etched, the nodeand fibril structure of the modified ePTFE was not destroyed orsubstantially altered.

Parallel with the molecular structure the wettability of the surfacechanged. For example, water drops were placed on ePTFE, PTFE and LDPEsurfaces. There was clear difference of the wettability on treatedsurfaces. The water drop on initial ePTFE could not be placed by thesteel cannula. The drop did not stay on the polymer surface but attachedto the needle. For measurements the drops fell down to the surface andwere measured then. For the treated ePTFE surface the drops were putfrom needle at usual procedure.

The contact angles of the water drops changed dramatically after plasmaimmersion ion implantation treatment. Low dose treatment induced higherhydrophilicty. At high doses the effect of etching and carbonizationcaused a decrease of wettability. This was observed both for PTFE andLDPE samples. The wetting measurement of ePTFE directly by the wettingangle technique is inadequate, because of the abnormal form of the drop.But the chemical structure of ePTFE and PTFE is quite similar and allowsthe transfer of the wetting measurement results of PTFE to ePTFEsamples.

FIG. 12 depicts contact angles of water on ePTFE (top), PTFE (middle),and LDPE (bottom) with plasma immersion ion implantation treatment withN⁺ at 20 keV and post-treatment as indicated. At post-treatment byacrylamide and HAES the wetting angles of PTFE and LDPE decreased tovery low values of 20-30 degrees. This wettability is similar as formetal surfaces. The treatment dose of 10¹⁴ and 10¹⁵ ions/cm² is enoughfor a good wettability. At high dose the effect of etching for PTFEsurface also was is observed for post-treated surfaces.

Influence of Silicon Deposition

In some experiments, unintendedly, some silicon deposition on polymersurface was observed. The reason was in previous experiments in thechamber with spattering of a silicon target and deposition of silicon onsubstrates, which had caused some remaining contamination. Thedeposition effect was observed by FTIR ATR spectra of LDPE. The strongintensity lines at 1090 and 1167 cm⁻¹ appeared in the spectra afterplasma immersion ion implantation. These lines are interpreted asvibrations of Si, SiO₂ and Si—C structures. In the field of 3400 cm⁻¹ awide line of Si—OH vibrations was observed. The intensity of these linesdid not depend on the dose of plasma immersion ion implantation andenergy of ions, but the amount of Si-containing structures depended ontime in plasma chamber. So, after plasma immersion ion implantation andsilicon deposition the surface of polymer contained a Si_(x)C_(y)O_(z)layer as combination of different structures.

Decreasing of plasma immersion ion implantation treatment time byincreasing of pulse repetition frequency did not could reduce theformation of the Si_(x)C_(y)O_(z) layer, but did not exclude it. Theeffect of Si-deposition was observed for different regimes of plasmaimmersion ion implantation, different types of ions, energies and pulsefrequencies.

However, the FTIR ATR spectra of ePTFE did not show any changes incomparison with untreated sample. The spectra had a low intensity ofpolytetrafluoro-macromolecule vibrations in comparison with PTFE film.The intensity of the macromolecule lines was more than 10 times lowerthan at samples without silicon deposition. The reason is the porestructure of ePTFE and the insufficient optical contact between the ATRcrystal and ePTFE surface for contrast spectra. In comparison with ePTFEafter pure plasma immersion ion implantation, at silicon deposition,there were no spectral changes corresponding to the carbonization of thesurface layer under ion bombardment. The color of the ePTFE changed lessthan at pure plasma immersion ion implantation. This indicates aprotective effect of the deposited layer on the ePTFE surface.

The contact angle measurements showed a strong influence of silicondeposition. The wetting angle depends on amount of Si—O—H groups andwettability can be very high. On the other side, at high doses of plasmaimmersion ion implantation the effect of interpenetration of Si atomsinto deep layers leads to increase of the contact angles.

Ageing of Plasma Immersion Ion Implantation Treated Surfaces

The ageing test was done for an analysis of stability of the modifiedsurfaces after plasma immersion ion implantation. The stability of thesurface was estimated by contact angle measurement of a water drop afterthermal ageing and after ageing and treatment by HAES. In the secondcase, not only wettability was tested, but the ability of the treatedsurface to preserve the chemical activity during ageing.

In the experiment, the samples of PTFE and ePTFE were treated by plasmaimmersion ion implantation at three different does, i.e., 10¹⁴ ions/cm²,10¹⁵ ions/cm² and 10¹⁶ ions/cm², as shown, and then treated by elevatedtemperature, which was used as model of the ageing process(time-temperature superposition). After thermal treatment the contactangles of PTFE remained mainly unchanged (FIG. 13A). The ageing kineticsshows that the hydrophilicty of treated PTFE can be preserved at leastfor several months.

At the second stage of experiments the plasma immersion ion implantationmodified samples after thermal treatment were treated with HAES and thewetting angles were measured. The contact angles increased with time ofageing (FIG. 13B). This indicates that the aged PTFE surface had lessactivity and less amount of HAES molecules reacted with the PTFEmodified layer. The chemical activity of modified PTFE decreases withtime. The chemical activity of the modified PTFE surface cannot be keptconstant over several months after plasma immersion ion implantationmodification.

The kinetics of the modified ePTFE samples had a more complex character.The wettability of an untreated sample decreased with time of thermalageing. Increase of wetting angle cannot be explained by oxidation ordestruction processes of the ePTFE macromolecules. Mainly the etchingeffect should lead to the increased hydrophobicity as releasing of wasteproducts from the surface layer of PTFE and appearance of an initialnon-defected ePTFE surface layer.

In the case of modified ePTFE samples, a decrease of wetting angles withtime of the ageing shows the structure transformation of the surfacelayer. The ageing of the ePTFE chemical activity shows more complexcharacter. Taken into account the complex dependence of wettability ofrough surfaces, the ageing kinetics by wetting angles have onlypreliminary character. These effects are depicted in FIGS. 14A and 14B,which show ePTFE contact angle as function of time in acceleratedageing. FIG. 14A is for ePTFE plasma immersion ion implantation with N⁺,implanted at the indicated doses of 10¹⁴ ions/cm², 10¹⁵ ions/cm² and10¹⁶ ions/cm². FIG. 14B is for ePTFE plasma immersion ion implantationwith N⁺, implanted at the indicated doses of 10¹⁴ ions/cm², 10¹⁵ions/cm² and 10¹⁶ ions/cm² and HAES post-treated.

Cell Culture

Step (1)

The purpose of this step mainly was to check, whether plasma immersionion implantation treatment of ePTFE causes the release of toxicby-products. For this no dose optimization or functionalization wasapplied. Cells used were GM7373 bovine aortic endothelial cells.

The metabolic activity of the cells on the samples was equivalent to 23%of total seeded cells on untreated ePTFE and 38% of total seeded cellson plasma immersion ion implantation treated ePTFE.

On the untreated ePTFE the initial polymer cells were small and round.Mainly they were between the ePTFE fibrils without direct contactthereto, only few adhered locally to the substrate. On treated ePTFEcells had an elongated body, which were aligned to the ePTFE fibrils. Acytoskeleton of polymerized f-actin was built up only on the plasmaimmersion ion implantation treated ePTFE, whereas on the untreated onethe actin in the cells formed unorganized aggregates.

The MEM elution test after three days did not show obvious morphologicdifferences of the cells with extracts from plasma immersion ionimplantation treated (N⁺ at 10¹⁶ ions/cm²) or untreated ePTFE or thenegative control. The 1 mM Cu²⁺, however caused severe cell damage.However, the inspection revealed a lot of fibrillar particles in theextract of irradiated ePTFE, what seemed to exceed the number ofparticles in the extract of initial ePTFE. The morphology of GM7373cells after three days exposure to extracts of ePTFE, Cu₂₊ as positivecontrol and a negative control are depicted in FIGS. 15A through 15D.

The exposure to medium, which was used in forming an extract generally,resulted in a slightly lower cell activity after three days than freshmedium. This behavior was dose dependent and independent, whetherinitial or plasma immersion ion implantation treated ePTFE was used. Thepositive control copper was toxic only in the highest concentration,whereas low concentrations even stimulated the metabolism as depicted inFIG. 16 in which cell count/activity of GM7373 cells after three daysexposure to with extracts of untreated ePTFE and nitrogen irradiated(10¹⁶ cm⁻²) ePTFE versus controls. Various dilutions with fresh mediumwere applied as indicated, Cu²⁺, respectively, was used in theconcentrations 1000, 250, 63, and 15.6 μmol/L.

Step (2)

The purpose of this step was a brief screening of different implantationregimes and regimes of functionalization.

Because of the generally low cell activity of cells on the substrates inthe step (1) experiments, the cell culture plastic Thermanox wasincluded here as a reference. On all ePTFE substrates the cell activityat day 1 was higher than on Thermanox. The cell activity was furtherclearly higher on the ion implanted ePTFE samples. Independent of thefunctionalization with acrylamide or with HAES, there seemed to be adose dependence of cell performance from the implantation dose, asindicated in FIG. 17. FIG. 17 depicts cell activity of GM7373 cellsafter 1 (white bars) and 3 days (gray bars) on the substrates asindicated, i.e., Thermanox, initial or untreated PTFE, PTFE treated ation concentrations of 10¹⁴ ions/cm², 10¹⁵ ions/cm², 10¹⁶ ions/cm², 10¹⁴and 10¹⁵ ions/cm² with acrylamide post treatment, and 10¹⁴, 10¹⁴ and10¹⁶ ions/cm² with HAES post treatment. The patterns indicate thecorresponding groups: the value of the third day was obtained byextended culture of the cells after the first Alamar Blue™ test; the100% reference is the same number of cells seeded on the bottom of acell culture well.

After the Alamar Blue™ test at day 1 the cells were further cultured andthe metabolic activity was determined again at day 3. For all types ofsample there the cell performance was dramatically decreased. At day 5almost no activity could be measured any more; fluorescent stain for thecell mitochondrial potential with JC-1 did not show any sign ofvitality. Stain for cell nuclei and f-actin also only showed remainingsof dead cells at day 5. It is more probable that this cell death is aneffect of the specific cell culture conditions with the Minusheetsupports and the repeated Alamar Blue™ test; both factors caused anunusual high oxygen partial pressure to the cells, which might induceoxidative stress. However, long-term investigations without intermittedmetabolic tests were not performed.

Step (3)

According to the findings in step (2) the highest dose treatment had thebest effect on cell adherence, therefore 10¹⁶ cm⁻² implanted sampleswith different functionalization were compared in more detail in thisstep.

In the same regime the Alamar Blue™ test was performed at day 1 and 3after seeding the cells. The drop in cell activity at day 3 wascompensated by normalizing the values to Thermanox as 100%.

Confirming the previous results, ion treatment improved the cellperformance. This was even more pronounced at day 3 than at day 1.Functionalization either with acrylamide or with HAES had the maineffect for initial adherence of the cells, but les for the longerperformance. Also the dose effect was more pronounced early afterseeding than at a later stage, as depicted in FIG. 18, which shows cellactivity normalized to Thermanox®D as 100%.

There was good cell spreading and the characteristic formation of acobblestone-like monolayer of the cells on Thermanox. The endothelialcells also adhere and spread on the plasma immersion ion implantationtreated and functionalized ePTFE, however, there the distinctarrangement of the cells was not present on the multifilament surface.Spreading of the cells also was seen on plasma immersion ionimplantation treated ePTFE without functionalization. On untreatedePTFE, cells mainly aggregated and did not adhere to the substrate.

CONCLUSIONS

Plasma immersion ion implantation was used for modification of ePTFE,PTFE and LDPE surfaces. The plasma immersion ion implantation treatmentwas done at different regimes, different gases, on two different plasmaimmersion ion implantation chambers. The modification of polymerstructures was observed by FTIR, XPS, wetting angle measurements. Thetesting showed the changes of molecular structure of surface layer,wettability changes, chemical activity changes and element contaminationchanges.

The effect of accelerated ageing of modified surfaces after plasmaimmersion ion implantation was observed. The wetting and chemicalactivity of PTFE and ePTFE surfaces were observed at ageing tests.

It was shown that such treated ePTFE does not release toxic degradationproducts to the medium, which would inhibit the growth or metabolism ofendothelial cells. In a static system the bovine aortic endothelial cellline GM7373 clearly adhered better on the ion treated ePTFE. There alsowas a higher metabolic activity of the cells on such treated samples,what can express both a higher activity of the individual cells orhigher absolute number of vital cells. The effect was slightly dosedependent. However, for high dose treatment the ePTFE fibrils alsotended to become more brittle. This was seen at the higher particlerelease in the extracts. Highly biostable particles of the size foundhere, in vivo tend to induce foreign body granulomas. Optimization ofthe treatment parameters and extensive washing processes have to beperformed in order to avoid this problem.

Treatment with higher energetic ions, ion implantation inducedcarburization of the surface, which prevented surface remodelling. Ionimplantation into polymers also lead to the formation of danglingbondings and free radicals in the polymer surface. Even though no effectwas seen here, these free radicals are potentially toxic for adherentcells. Therefore these bondings were saturated with the highlyhydrophilic acrylamide. The substance by this process covalently bondsto the polymer surface and free monomers of the toxic substance werewashed out. In the experiments described above a better performance ofacrylamide treated ion implanted ePTFE was shown. Alternatively toacrylamide, the modified polysaccharide hydroxyethyl starch or HAES wasalso used to saturate the free radicals on the polymer surface. HAES isin clinical application as plasma expander for the treatment ofhypovolaemia and shock. Due to the substitution it is better soluble inwater than ordinary starch and more resistant against degradation. Insolution it tends to inhibit haemostatic processes by interaction withclotting factors and blood platelets. Endothelial cells also internalizethe free form by pinocytosis and eliminate only about 50% of it. Theexpression of activation markers is not modulated by this, but HAESdirectly inhibits the interaction with polymorphonuclear neutrophils.

In this study, HAES may be also seen more as a model substance, whichcan be used to functionalize the surface. It can be either substitutedwith glucosaminogycans like heparin, or direct modification of thehydroxyethyl group e.g. with the before mentioned elastin or peptideswould be possible. To our knowledge, this would be a new approach tofunctionalize the ePTFE surface for a specific and strong adherence ofendothelial cells.

The invention being thus described, it will now be evident to thoseskilled in the art that the same may be varied in many ways. Suchvariations are not to be regarded as a departure from the spirit andscope of the invention and all such modifications are intended to beincluded within the scope of the following claims.

1. An implantable medical device comprising: ePTFE having a surfacemodified by plasma immersion ion implantation; and polysaccharidehydroxyethyl starch groups covalently bonded to the modified surface. 2.The implantable medical device of claim 1, further comprising abioactive agent bonded to the polysaccharide hydroxyethyl starch groups.3. The implantable medical device of claim 2, further comprising abiodegradable spacer molecule bonded to the modified surface or to thepolysaccharide hydroxyethyl starch groups.
 4. The implantable medicaldevice of claim 2, wherein the bioactive agent is selected from thegroup consisting of anti-thrombogenic agents, anti-proliferative agents,anti-inflammatory agents, antineoplastic/antiproliferative/anti-mitoticagents, anti-coagulants, vascular cell growth promoters, vascular cellgrowth inhibitors, cholesterol-lowering agents; vasodilating agents;agents which interfere with endogenous vasoactive mechanisms, celladhesion factors; and combinations thereof.
 5. The implantable medicaldevice of claim 1, wherein the device is a graft or a stent-graft.
 6. Animplantable medical device comprising: ePTFE having a surface modifiedby plasma immersion ion implantation; and hydrophilic acrylamide groupscovalently bonded to the modified surface.
 7. The implantable medicaldevice of claim 6, further comprising a bioactive agent bonded to thehydrophilic acrylamide groups.
 8. The implantable medical device ofclaim 6, further comprising a biodegradable spacer molecule bonded tothe modified surface or to the hydrophilic acrylamide groups.
 9. Theimplantable medical device of claim 7, wherein the bioactive agentcomprises anti-thrombogenic agents, anti-proliferative agents,anti-inflammatory agents, antineoplastic/antiproliferative/anti-mitoticagents, anti-coagulants, vascular cell growth promoters, vascular cellgrowth inhibitors, cholesterol-lowering agents; vasodilating agents;agents which interfere with endogenous vasoactive mechanisms; celladhesion factors; and combinations thereof.
 10. The implantable medicaldevice of claim 6, wherein the device is a graft or a stent-graft. 11.An implanted, surface modified ePTFE graft, comprising: ePTFE having anode and fibril structure and having a carburized surface formed byplasma immersion ion implantation without destroying the node and fibrilstructure and having cellular material attached to the fibrils orattached to functional groups covalently bonded to the fibrils andsubstantially covering and/or filling the nodes.